Effectiveness and safety of auricular acupoint bloodletting in treatment of insomnia: an assessor-blinded pilot randomized controlled trial.

OBJECTIVE: To evaluate the effectiveness and safety of auricular acupoint bloodletting in treatment of insomnia METHODS: Participants (n = 60) with insomnia were randomized into two groups to receive treatment of auricular acupoint bloodletting: low frequency group, 1 times/week for five weeks (n = 30); high frequency group, 2times/week for two weeks (n = 30). The following outcomes were measured blindly at baseline, after first treatment, 4 weeks, and 8 weeks: Pittsburgh sleep quality index scale (PSQI). RESULTS: The groups were balanced at baseline for insomnia and demographic characteristics. There were no significant differences between the groups in terms of any of the outcomes, at the first follow-up time point. However, the therapeutic effect of LFG (once per week) is obviously lower than that of HFG (twice per week). In addition, there was no significant difference in the side effects between the two groups. CONCLUSION: The treatment of insomnia with different frequencies of auricular acupoint bloodletting is effective and has less side effects. More reasonable treatment frequencies are worth further study.

Hypoxic preconditioning up-regulates glucose transport activity and glucose transporter (GLUT1 and GLUT3) gene expression after acute anoxic exposure in the cultured rat hippocampal neurons and astrocytes.

Hypoxic preconditioning has been shown to increase the hypoxic tolerance of brain neurons. However, the mechanism underlying the increased hypoxic tolerance has not been well elucidated. Since anaerobic glycolysis is the only pathway for a vertebrate cell to produce energy under anoxic conditions, which needs a large amount of glucose, we hypothesize that glucose transport, the rate-limiting step for glucose metabolism, plays a critical role in the hypoxic tolerance induced by hypoxic preconditioning. In this study, the effects of hypoxic preconditioning on glucose transport activity and the gene expression of two major forms of glucose transporters (GLUT1 and GLUT3) in the brain were investigated in cultured rat hippocampal neurons and astrocytes. The neuronal and astroglial cultures were preconditioned for 6 days by intermittently exposing the cells to sublethal hypoxic gas mixture (1% O2/10% CO2/89% N2) for 20 min each day. 24 h after the last hypoxic exposure, the cells were exposed to a lethal anoxic gas mixture (10% CO2/90% N2) for 6 h and the uptake rate of [3H] 2-deoxyglucose (2-DG) and the levels of GLUT1 and GLUT3 glucose transporter mRNAs in the cells were examined immediately after anoxic exposure. The neurons and astrocytes preconditioned with hypoxia showed higher 2-DG uptake rates than the non-preconditioned cells. Compatible with the change in 2-DG uptake, hypoxic preconditioning also increased GLUT1 mRNA levels in the astrocytes and GLUT1 and GLUT3 mRNA levels in the neurons. The neurons preconditioned by hypoxia displayed increased anoxic tolerance. However, when glucose uptake in the neurons was blocked by cytochalasin B, the anoxic tolerance was almost abolished. These results suggest that glucose transport is critical to neuronal survival during anoxic exposure and the increased glucose transport activity is probably one of the important mechanisms for the enhanced hypoxic tolerance induced by hypoxic preconditioning.

Underlying mechanism of hypoxic preconditioning decreasing apoptosis induced by anoxia in cultured hippocampal neurons.

It is known that hypoxic preconditioning (HP, a brief period of sublethal hypoxia) provides neuroprotection against subsequent severe anoxia, but the mechanisms of this increased tolerance have not been fully elucidated. A hypoxic preconditioning model was established by exposing a 4-day hippocampal culture to 1% O(2) for 20 min/day for 8 days. The preconditioning significantly decreased the number of apoptotic neurons at reoxygenation 24 h after 4 h of severe anoxia (0% O(2)). Further study demonstrated that the degradation of mitochondrial membrane potential (MMP) was greatly inhibited and the expression of B-cell lymphoma protein-2 (Bcl-2) was increased considerably after severe anoxia in the HP groups. These results indicate that the increased anoxic tolerance, which is induced by HP in cultured hippocampal cells, may be correlated with Bcl-2 overexpression and enhanced stability of MMP, which ultimately reduces apoptosis 24 h after reoxygenation.

Involvement of increased stability of mitochondrial membrane potential and overexpression of Bcl-2 in enhanced anoxic tolerance induced by hypoxic preconditioning in cultured hypothalamic neurons.

The effects of hypoxic preconditioning (HP) on changes in mitochondrial membrane potential (MMP) and Bcl-2 expression in cultured hypothalamic neurons after severe anoxia were investigated. In the HP group, hypothalamic neurons, after a 4-day culture, were preconditioned daily under a hypoxic condition (1% O(2), 10 min) for 8 days; subsequently, the HP neurons and those in the control group (similarly cultured, but without HP) were exposed to 6 h of severe anoxia (0% O(2)). The preconditioned neurons had a higher survival rate and a lower lactate dehydrogenase leakage, compared with the control group. Although HP did not prevent the degradation of MMP during severe hypoxia, preconditioned neurons exhibited a higher level of MMP than that of the control group. Increased expression of Bcl-2 was also observed in the preconditioned hypothalamic neurons. These results suggest that HP enhances the hypoxic tolerance of hypothalamic neurons, and the underlying mechanisms may be related to the increased stability of MMP and the overexpression of Bcl-2 induced by HP.

[Effect of CoCl2 pretreatment on Na + and K+ currents of the rat hippocampal neurons after acute hypoxia].

AIM: To study effect of CoCl2 pretreatment on the voltage-gated Na+ and K+ currents of the rat hippocampal neurons after acute hypoxia. METHODS: Primarily cultured hippocampal neurons were divided into CoCl2 pretreated and non-pretreated groups. Patch clamp whole cell recording technique was used to examine Na+ and K+ currents of the hippocampal neurons. RESULTS: After acute hypoxia, I(Na) and I(K) of the hippocampal neurons were significantly decreased and the threshold of I(Na) was right-shifted. Pretreatment of the neurons with CoCl2 inhibited the reduction of I(Na) and I(K). CONCLUSION: CcCl2 pretreatment alleviates the acute hypoxia-induced changes of I(Na) and I(K), which may be one of the mechanisms for the protective effect of CoCl2 on neurons.

[Protective effect of GABA against hypoxic injury in rat hippocampal slices].

AIM: To study the mechanism of protective effect of GABA against hypoxic injury in rat hippocampal slices. METHODS: The hippocampal slices from adult rats and extracellular recording technique were used to observe the effect of GABA on the evoked population spikes in rat hippocampal slices after hypoxia in vitro. RESULTS: GABA can significantly delay the disappearance of PV, but have no effect on PS. When the receptor antagonist of GABA (bicuculline) and the inhibitor of Cl- channel (NPPB) were given, the protect effect could be suppressed. CONCLUSION: GABA increases hypoxic tolerance of hippocampal slices. The mechanism of the effect of GABA may be involved in the elevation of chloride influx through GABA receptor.

[CoCl2-induced enhancement of glucose transport activity in mediating hypoxic tolerance in cultured hippocampal neurons].

The effect of CoCl(2) pretreatment on glucose transport activity of cultured newborn rat hippocampal neurons and its role in neuronal hypoxic tolerance were observed. The results showed that the 2-deoxy-D-[1-(3)H ]glucose uptake rate and the mRNA expressions of glucose transporters (GLUT1 and GLUT3) in the hippocampal neurons were significantly increased after a 24-hour pretreatment with CoCl(2). The cell injury induced by 6-hour or 8-hour hypoxic exposure was also greatly reduced by CoCl(2) pretreatment. The protective effect of CoCl(2) on the neurons was largely abolished by cytochalasin B, a specific inhibitor of glucose transporters. The results suggest that CoCl(2) can increase mRNA expressions of GLUT1 and GLUT3 and glucose transporter activity of the neurons, which may be an important mechanism for the increased tolerance of the neurons to hypoxia.

[Effects of rhIL-6 on Bcl-2 and Bax expression and apoptosis after anoxia-reoxygenation in cultured rat hippocampal neurons].

The purpose of the present study was to determine the effects of recombinant human interleukin-6 (rhIL-6) on the Bcl-2 and Bax expression and apoptosis after anoxia-reoxygenation in cultured rat hippocampal neurons. The control and rhIL-6 treated hippocampal neurons cultured for 12 d were exposed to anoxia environment (90% N2+10% CO2) for 2 and 4 h and then were reoxygenated for 24 and 72 h. The expression of Bcl-2 and Bax was revealed immunocytochemically using the antiserum against Bcl-2 and Bax. The apoptosis was examined by the terminal deoxynucleotidyl transferase-mediated biotinylated deoxyuridine triphosphate nickel end labeling (TUNEL) method and flow cytometric analysis. The results showed that in cultured hippocampal neurons the Bcl-2 expression decreased while Bax expression and the percentage of apoptotic neurons increased after anoxia-reoxygenation compared with those before anoxia. In comparison with the control, after anoxia-reoxygenation the Bcl-2 expression in hippocampal neurons was higher than that in rhIL-6 group; however the Bax expression and the percentage of the apoptosis were decreased in rhIL-6 group. It is suggested that rhIL-6 may play a role in protecting neurons from the damage induced by anoxia-reoxygenation.

[Effects of hypoxic-preconditioning on anoxic-tolerance and Jun expression in cultured rat hippocampal neurons].

AIM: To study the effects of hypoxic preconditioning on anoxic tolerance and Jun expression in cultured rat hippocampal neurons after anoxia/reoxygenation. METHODS: 12 day cultured hippocampal neurons in control and hypoxic preconditioning group were exposed to anoxic environment (0.90L/L N2 + 0.10 L/L CO2) for 4 h, and then reoxygenated for either 24 h or 72 h. The neurons were immunocytochemically stained using the antiserum against Jun. The number of survival neurons and the percentage of Jun expressing neurons were investigated. RESULTS: The percentage of Jun expressing neurons induced by anoxia in hypoxic-preconditioning group was significantly less than that in control group. The number of survival neurons was more in the hypoxic-preconditioning group than that in control group after anoxic reoxygenation. CONCLUSION: Hypoxic-preconditioning can induce the development of anoxic-tolerance in cultured hippocampal neurons. The decrease in Jun expressing neurons in hippocampus may be an adaptive reaction to acute anoxia.

[Relationship between enhanced anoxic tolerance induced by hypoxic preconditioning and Na+, K+ currents in cultured hypothalamic cells].

AIM: To investigate the relationship between enhanced anoxic tolerance induced by hypoxic preconditioning and Na+, K+ currents. METHODS: After hypoxic preconditioning and acute anoxia the I(Na), I(K) were measured in cultured hypothalamic cells by patch-clamp whole cell recording technique. RESULTS: The amplification of Na+ currents did not been significantly changed, but the amplification of K+ currents was in hypoxic preconditioning neurons; acute anoxia lead to the inhibition of Na+, K+ currents in the two groups, while Na+, K+ currents in non-preconditioned control group were inhibited severity than hypoxic preconditioning group. CONCLUSION: It is presumed enhanced anoxia tolerance induced by hypoxic preconditioning may be related to the opening of K+ channels.

[Effects of anoxia/reoxygenation on Fos and Jun expression and apoptosis in cultured rat hippocampal neurons].

AIM: To investigate the effects of anoxia/reoxygenation on Fos and Jun expression and apoptosis in cultured rat hippocampal neurons. METHODS: The hippocampal neurons cultured for 12 d were exposed to anoxia environment (90% N2 + 10% CO2) for 4 h and then reoxygenated for 24 h and 72 h. The neurons were immunocytochemically stained using the antiserum against Fos and Jun, and the apoptosis were detected by using the terminal deoxynucleotidyl transferase mediated biotinylated deoxyuridine triphosphate nickel end labeling (TUNEL) method and flow cytometric analysis. RESULTS: The percentage of Fos and Jun positive neurons and apoptosis neurons in cultured hippocampal neurons after anoxia/reoxygenation increased than those in control. CONCLUSION: The occurrence of neurons apoptosis is related to the increase in Fos and Jun expression in cultured hippocampal neurons after anoxia/reoxygenation.